D 2 Dopamine Receptors on Bovine Chromaffin Cell Membranes: Identification and Characterization by [ 3 H] N ‐Methylspiperone Binding

Although dopamine‐containing cells are known to be present in sympathetic ganglia, the site of action and the role of dopamine in ganglion function remain obscure. In the present work, we evaluated the interaction of dopamine receptor ligands with particulate membrane fractions from bovine chromaffin cells and adrenal medullary homogenates using the D 2 dopamine receptor radioligand [ 3 H] N ‐methylspiperone ([ 3 H]NMSP). Scatchard analysis of [ 3 H]NMSP saturation experiments revealed a B max of 24.1 ± 1.6 fmol/mg of protein and a K D of 0.23 ± 0.03 n M in the particulate fraction from adrenal medulla homogenates and a B max of 26.5 ± 2.7 fmol/mg of membrane protein and a K D of 0.25 ± 0.02 n M in the particulate fraction prepared from isolated adrenal chromaffin cells. There were ∼ 1,000 receptors/cell. There were no detectable levels of specific [ 3 H]NMSP binding in the particulates prepared from adrenal cortical or capsular homogenates. Competition studies with the nonradioactive D 2 receptor antagonists spiperone, chlorpromazine, and (‐)‐sulpiride revealed K 1 values of 0.28, 21, and 196 nM, respectively. The (+) isomer of butaclamol displayed a 604‐fold higher affinity than the (‐) isomer. Competition studies with the dopamine receptor agonists dopamine and apomorphine revealed affinities of 3,960 and 417 n M , respectively. A correlation coefficient of 0.96 was obtained in studies comparing the potencies of drugs in inhibiting specific [ 3 H]NMSP binding in bovine adrenal medullary homogenates and in inhibiting specific [ 3 H]NMSP binding to brain D 2 dopamine receptors. In summary, radiolabeling studies using [ 3 H]NMSP have revealed the presence of D 2 dopamine receptors on bovine adrenal chromaffin cells. Dopamine may modulate adrenal medullary function, and possibly sympathetic ganglion function, through D 2 dopamine receptors.